Cell migrations are a characteristic of multicellular life, both during development and malignant or, pathological states. A molecularly complex extracellular landscape plays a crucial role in directing migrating cells. One of the most complex structures in the extracellular space is heparan sulfate proteoglycans (HSPGs) that have been shown to be essential for cell-cell signaling. Here we propose to study the function of heparan sulfate proteoglycans in cellular migration using stereotypic cell migrations in the small nematode C. elegans which require HSPGs for function. To this end we are using an integrated approach that combines biochemical studies with genetic approaches to directly correlate protein and sugar structure of HSPGs with their function during cell migration. Our project will compare distinct cell migrations that we know already employ distinct molecular mechanisms, including distinct HS sugar structures. Lastly, we will define the global HS landscape that migrating cells encounter. To this end we will use a novel tool we have developed that allows the direct visualization of HS motifs in living animals. Our studies will allow correlating HS structure with cellular and subcellular localization. In summary, the proposed studies will provide insight into the cell-specific and sugar-dependent mechanisms that migrating cells use to interpret the extracellular space. Understanding fundamental aspects of cell-specific interactions of migrating cells may ultimately allow the cell-specific intervention during diseased states.